Submitted by Victor Alexandre Zonaro (victor.zonaro@gmail.com) on 2017-01-10T14:22:50Z No. of bitstreams: 1 Dissertação - Victor Alexadre Zonaro.pdf: 4719611 bytes, checksum: 075493217f07f41d58c2e71196b60eea (MD5) / Approved for entry into archive by LUIZA DE MENEZES ROMANETTO (luizamenezes@reitoria.unesp.br) on 2017-01-13T17:16:22Z (GMT) No. of bitstreams: 1 zonaro_va_me_araiq.pdf: 4719611 bytes, checksum: 075493217f07f41d58c2e71196b60eea (MD5) / Made available in DSpace on 2017-01-13T17:16:22Z (GMT). No. of bitstreams: 1 zonaro_va_me_araiq.pdf: 4719611 bytes, checksum: 075493217f07f41d58c2e71196b60eea (MD5) Previous issue date: 2016-12-20 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / As 5,6-diidro-2-pironas 6-substituídas, também conhecidas como estirilpironas, são uma importante classe de metabólitos secundários presentes em plantas. São substâncias biologicamente ativas, apresentando como, por exemplo, atividade anticâncer, antioxidante, antifúngica e antiviral. O gênero Cryptocarya, pertence à família Lauraceae e apresenta diversas estirilpironas em suas mais diversas partes: folhas, cascas, sementes e raízes. Foram selecionadas para análise as espécies C. mandioccana, C. moschata e C. botelhensis. Foram utilizadas apenas as folhas, por apresentarem facilidade para coleta e abundância. Este trabalho tem como objetivo a identificação das estirilpironas presentes em três espécies brasileiras de Cryptocarya com o uso das técnicas HPLC-DAD-MS. Foi desenvolvido método cromatográfico para análise do extrato hidroalcoólico das folhas de espécies de Cryptocarya por HPLC-DAD-MS utilizando etanol como fase orgânica na fase móvel. Com os espectros no UV foi possível identificar que as classes de metabólitos presentes nas amostras eram alcaloides, flavonoides e estirilpironas. Utilizando os dados de MS e MS/MS, foi possível a caracterização de estirilpironas presentes nos extratos, assim como a sugestão de suas fragmentações por espectrometria de massas, além da identificação dos íons m/z 163 e m/z 189 característicos da fragmentação das estirilpironas. Foi detectado o íon m/z 423 no extrato de C. moschata, referente a um possível dímero da goniotalamina, sem relato anterior para as espécies de Cryptocarya. Com o auxilio de pironas purificadas obtidas pelo grupo, foi possível a comparação de seus tempos de retenção com os dados obtidos a partir dos extratos de folhas das espécies de Cryptocarya ajudando na identificação das substâncias presentes. Além das estirilpironas, foram identificados os alcalóides menisperina e xantoplanina nas três espécies estudadas. Nota-se uma grande diferença na quantidade de metabólitos entre as espécies analisadas, sendo que a C. mandioccana é a mais rica em estirilpironas. / The 6-substituted 5,6-dihydro-2-pyrones, also known as styrylpyrones, are an important class of secondary metabolites present in plants. They are biologically active substances, presenting, for example, anticancer, antioxidant, antifungal and antiviral activity. The genus Cryptocarya belongs to the family Lauraceae and presents several styrylpyrones in its most diverse parts: leaves, barks, seeds and roots. The species C. mandioccana, C. moschata and C. botelhensiswere selected for analysis. We chose to use only leaves, because they are easy to collect and abundant. The objective of this work is to identify the styrylpyrones present in three Brazilian Cryptocarya species using the HPLC-DAD-MS techniques. A chromatographic method was developed to analyze the hydroalcoholic extract from leavesCryptocarya species by HPLC-DAD-MS using ethanol as the organic solvent in the mobile phase. With the UV spectra were possible to identify the classes of metabolites present in the samples as alkaloids, flavonoids and styrylpyrones. Addtitionaly, with the MS and MS2 data, was possible to characterize the styrylpyrones present in the extracts, as well as the suggestion of their fragments by mass spectrometry, in addition to the identification of íons m/z 163 and m/z 189 characteristics of the fragmentation of styrylpirones. The m/z 423 ion detected in C. moschata extract was tentatively attributed to a goniotalamine dimer, with no previuous reports for Cryptocarya species. With the help of purified pirones obtained by the group, it was possible to compare their retention times with the data obtained from leaf extracts of the Cryptocarya species, helping to identify the substances present. Besides the styrylpyrones, the alkaloids menisperin and xantoplanin were also identified in the three species studied. There is a great difference in the amount of metabolites among the analyzed species, with C. mandioccana being the richest in styrylpyrones.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-10-26T20:21:40Z No. of bitstreams: 1 MoniqueGomesDantas_DISSERT.pdf: 3471537 bytes, checksum: a73f34ecbaa2b638849e2cfdb0b85408 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-12-20T18:08:05Z (GMT) No. of bitstreams: 1 MoniqueGomesDantas_DISSERT.pdf: 3471537 bytes, checksum: a73f34ecbaa2b638849e2cfdb0b85408 (MD5) / Made available in DSpace on 2016-12-20T18:08:05Z (GMT). No. of bitstreams: 1 MoniqueGomesDantas_DISSERT.pdf: 3471537 bytes, checksum: a73f34ecbaa2b638849e2cfdb0b85408 (MD5) Previous issue date: 2015-02-06 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) / Guanilhidrazonas s?o uma classe de subst?ncias amplamente estudadas por apresentar grande potencial biol?gico. A an?lise de f?rmacos e medicamentos ? ferramenta importante para garantir qualidade, seguran?a e efic?cia aos novos medicamentos. Nesse estudo foram avaliadas as guanilhidrazonas sint?ticas WE005 (3,4-dimetoxibenzalde?doguanilhidrazona), WE015 (benzalde?doguanilhidrazona) e WE016 (metil-4-formilbenzoatoguanilhidrazona) com o objetivo de caracterizar, desenvolver e validar m?todo anal?tico utilizando t?cnicas t?rmicas (DSC e TG), espectrosc?pica (FTIR), microsc?pica (MEV) e cromatogr?fica (CLUE/DAD). Na caracteriza??o por DSC e TG foram utilizadas as seguintes raz?es de aquecimento: 2,5; 5,0; 10 e 20 ?C/min em atmosfera de nitrog?nio (50 mL/min) at? 500?C (DSC) e 900?C (TG). A an?lise na regi?o do infravermelho m?dio das mol?culas foi realizada a temperatura ambiente e na faixa de fus?o. Os espectros foram comparados atrav?s da correla??o de Pearson utilizando o algoritmo ad hoc. O estudo cin?tico foi feito atrav?s do m?todo de Ozawa. O planejamento fatorial investigou a influ?ncia do comprimento da coluna, fluxo e propor??o de fase m?vel sobre o tempo de reten??o, fator de cauda, resolu??o e n?mero de pratos te?ricos. As t?cnicas t?rmicas foram capazes de caracterizar as mol?culas atrav?s de suas transi??es de fase e etapas na curva termogravim?trica, informando ainda sobre a estabilidade t?rmica, com temperatura inicial de decomposi??o em torno de 240 ?C. O estudo cin?tico mostrou que todas as mol?culas apresentam ordem zero e que a amostra WE016 apresentou maior energia de ativa??o. Os espectros de infravermelho de acordo com a correla??o de Pearson apresentaram mudan?as significativas entre a temperatura ambiente e o espectro da faixa de fus?o. O planejamento fatorial atrav?s dos gr?ficos de superf?cie-resposta e Pareto revelou que a vari?vel de maior influ?ncia sobre todas as vari?veis dependentes foi o comprimento da coluna. O melhor m?todo para a separa??o das guanilhidrazonas deste estudo foi: coluna C18 (50 mm x 2 mm t.p. 2.2 ?m), fase m?vel MeOH:H2O:TEA 40:60:0,1, pH 3,5 ajustado com ?cido ac?tico; fluxo de 0,2 mL/min, temperatura do forno 30 ?C. A concentra??o final das guanilhidrazonas foi de 30 ?g/mL e foram detectadas simultaneamente atrav?s do comprimento de onda de 290 nm. Um m?todo r?pido foi desenvolvido para separar as guanilhidrazonas WE005, WE015 E WE016 por CLUE/DAD. Planejamento fatorial foi uma ferramenta ?til para o desenvolvimento racional do m?todo. / Guanilhidrazonas substances are widely studied for presenting great biological potential, especially antitumor activity, antimicrobial, antimalarial, antifungal and anti-hypertensive. This study assessed the synthetic guanylhydrazones WE005 ((2-[(3,4-dimethoxyphenyl)methylene]-hydrazinecarboximidamide),WE015 (2(phenylmethylene)-hydrazinecarboximidamide) and WE016 (4-[[2-(aminoiminomethyl) hydrazinylidene]methyl]-benzoic acid methyl ester) in order to characterize and develop analytical method using thermal techniques (DSC and TGA), spectroscopic (FTIR ) and microscopic (SEM) and chromatographic (UHPLC / DAD). The characterization by DSC and TG were used the following heating rates: 2.5; 5.0; 10 and 20 ? C / min in a nitrogen atmosphere (50 ml / min) to 500 ? C (DSC) and 900 ? C (TG). The kinetic study was done by Ozawa method. The analysis in the mid-infrared region of molecules was performed at room temperature and the melting temperature. The spectra were compared using Pearson's correlation using ad hoc algorithm. For the development of the analytical method was used factorial design. Thermal techniques have been able to characterize the molecules through their phase transitions and steps in the thermogravimetric curve, and telling about the thermal stability, with an initial decomposition temperature of about 240 ? C. The kinetic study showed that the speed of the thermal decomposition of molecules has zero order and that the WE016 sample showed higher activation energy. Infrared spectra according to the Pearson correlation showed significant changes between the room temperature and the melting range of the spectrum. The factorial design through surface-response graphs and Pareto showed that the most influential variable on all dependent variables was the length of the column. The best method for the separation of guanylhydrazones this study was: C18 column (50 mm x 2 mm tp 2.2 microns), mobile phase MeOH: H2 O: TEA 40: 60: 0.1, pH 3.5 adjusted with acetic acid; flow rate of 0.2 ml / min, oven temperature 30 ? C. The final concentration of guanylhydrazones was 30 mg / mL and were detected simultaneously by a wavelength of 290 nm. A rapid method was developed to separate guanylhydrazones WE005, WE015 and WE016 by CLUE / DAD. Factorial design was a useful tool for the rational development of the method.

Submitted by Marlene Santos (marlene.bc.ufg@gmail.com) on 2018-07-09T16:12:59Z No. of bitstreams: 2 Tese- Maysa Aparecida de Oliveira - 2013.pdf: 2303389 bytes, checksum: d6b1b0186001f327c5e50809b8d401fb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-07-10T11:10:17Z (GMT) No. of bitstreams: 2 Tese- Maysa Aparecida de Oliveira - 2013.pdf: 2303389 bytes, checksum: d6b1b0186001f327c5e50809b8d401fb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-07-10T11:10:17Z (GMT). No. of bitstreams: 2 Tese- Maysa Aparecida de Oliveira - 2013.pdf: 2303389 bytes, checksum: d6b1b0186001f327c5e50809b8d401fb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2013-07-16 / A simple and accurate stability-indicating high-performance liquid chromatography (HPLC-DAD) method was developed to measure nifedipine (NIF) in plasma in the presence of its degradation products. The chromatographic separation was performed on a C18 column using H3PO4 0.01%:CH3CN:CH3OH (60:20:20, v/v/v) as the mobile phase and a 1.0ml/min flow rate. The analytical validation was performed according to FDA, EMA and ANVISA (Brazilian Health Surveillance Agency) guidelines and was linear between 100.0 and 2000.0 ng/ml, precise (1.9 to 11.3%) and accurate (86.0 to 113.1%). Under the evaluated conditions, NIF in plasma samples were stable after processing and freeze-thaw cycles. The average liquid-liquid extraction recovery was 101.3 ± 5.4%. After validation, this method was applied to evaluate the influence of nitrosophenylpyridine (NO-NIF) in the pharmacokinetic of NIF in plasma of Wistar rats. This method was also validated for quality control of NO-NIF obtained after photodegradation of NIF in photostability chamber. The method showed selectivity, linearity (0.4 to 2.4mg/ml), as well as precision and accuracy. Nitrophenylpyridine (NINIF) was synthesized from the NIF. These degradation products were characterized by NMR and mass spectroscopy. In addition, a breakdown product of NO-NIF due to its contact with plasma was identified and characterized. / Um método simples, rápido e preciso por cromatografia líquida de alta eficiência (HPLC-DAD) foi desenvolvido para determinação de nifedipino (NIF) na presença de seus produtos de degradação em amostras de plasma. A separação cromatográfica foi realizada em coluna C18 empregando H3PO4 0,01%:CH3CN:CH3OH (60:20:20 v/v/v) como fase móvel e vazão de 1,0mL/min. A validação procedeu-se segundo as recomendações dos guias editados pela ANVISA, EMA e FDA e apresentou linearidade no intervalo de 100,0 a 2000,0ng/mL com precisão (1,9 a 11,3%) e exatidão 86,0 a 113,1%) adequadas. Nas condições avaliadas, as amostras de NIF mostraram-se estáveis tanto em plasma como após processamento e ciclos de congelamento e descongelamento. A eficiência do método de extração líquido-líquido demonstrou recuperação média de 101,3 ± 5,4%. Após a etapa de validação, o método foi aplicado em estudos farmacocinéticos para avaliação da influência do nitrosofenilpiridino (NO-NIF) na farmacocinética do NIF. Este método também foi validado para o controle de qualidade do NO-NIF obtido após fotodegradação do NIF em câmara de fotoestabilidade e apresentou seletividade, linearidade (0,4 a 2,4μg/mL), assim como precisão e exatidão. Além da obtenção do NO-NIF por fotodegradação, nitrofenilpiridino (NI-NIF) foi sintetizado a partir do NIF. Esses produtos de degradação foram caracterizados por RMN e espectrometria de massas. Além disso, um produto de degradação do NO-NIF decorrente do seu contato com plasma foi identificado e caracterizado.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / Agriculture is one of the main economic activities of the Brazil, possessing the culture of coffee as their main primary commodity. In order to achieve high levels of production and prevent the loss of crops, farm inputs such as pesticides are used in large quantities. This type of practice and the increased public concern consumer are motivating scientific research about these topics, with the aim of ensuring food safety of products supplied for consumption. However, the literature does not report methodologies for the determination of pesticide residues in roasted coffee, with the methods of extraction and analysis proposed, which according to bibliographic data present residues of these substances even after roasting. In this context, the present study sought to develop an analytical methodology to determine carbofuran, cypermethrin, chlorpyrifos, clothianidin, disulfoton, endosulfan, spirodiclofen, haloxyfop, imidacloprid, tebuconazole, triadimenol and triadimefom in roasted coffee using solid liquid extraction method by sonication with clean-up step through liquid liquid extraction (LLE) and instrumental analysis for ultra performance liquid chromatography tandem mass spectrometry detector (LC-MS/MS). For both, the optimization of chromatographic conditions and assessing the best method of clean-up was carried out. With that, the conditions for better quantitative response was obtained with 0.5 g of roasted coffee, 5 mL extraction solvent by sonication, 1 mL of water and 1 mL of dichloromethane in the clean-up by LLE. During the validation procedure in LC-MS/MS positive matrix effect was observed for all analytes except of cypermethrin in which there was suppression of the signal by array, causing, prepared in the curve-based extract, recoveries between 74.2%-102.4% standard relative deviation (SRD) between 0.6% and 10.2%, in levels concentration assessed in the process, Furthermore, the linearity obtained was in the range of 0.9979 to 0.9998 for pesticides , carbofuran, disulfoton, imidacloprid, clothianidin tebuconazole, triadimenol, triadimefom and ensuring the efficiency of the methodology. / A agricultura é uma das principais atividades econômicas do Brasil, possuindo a cultura do café como sua principal commodity primária. A fim de atingir altos níveis de produção e evitar a perda das safras, insumos agrícolas como os agrotóxicos são utilizados em grandes quantidades. Este tipo de prática e o aumento da preocupação do público consumidor estão motivando investigações científicas a respeito destes tópicos, com o intuito de garantir a segurança alimentar dos produtos fornecidos para o consumo. No entanto, a literatura não relata metodologias para a determinação de resíduos de agrotóxicos no café torrado, com os métodos de extração e análise propostos, que segundo dados bibliográficos apresentam resíduos destas substâncias mesmo após a torrefação. Neste contexto, o presente trabalho buscou desenvolver uma metodologia analítica para determinar carbofurano, cipermetrina, clorpirifós, clotianidina, dissulfotom, endosulfan, espirodiclofeno, haloxifope, imidacloprido, tebuconazol, triadimefom e triadimenol em café torrado empregando método de extração sólido líquido por sonicação com etapa de clean-up por meio de extração líquido líquido (LLE) e análise instrumental por cromatografia líquida de ultra eficiência acoplada a detector por espectrometria de massas (LC-MS/MS). Para tanto, a otimização das condições cromatográficas e a avaliação do melhor método de clean-up foram realizadas. Com isso, as condições de melhor resposta quantitativa foi obtida com 0,5 g de café torrado, 5 mL de solvente de extração por sonicação, 1 mL de água e 1 mL de diclorometano na etapa de clean-up por LLE. Durante o procedimento de validação no LC-MS/MS foi observado efeito matriz positivo para todos os analitos com exceção da cipermetrina na qual houve supressão do sinal pela matriz, ocasionando, com base na curva preparada no extrato, recuperações entre 74,2% - 102,4%, com desvio padrão relativo entre 0,6% e 10,2%, nos níveis de concentração avaliados no processo, além disso, a linearidade obtida foi na faixa de 0,9979 a 0,9998 para os agrotóxicos, carbofurano, clotianidina dissulfotom, imidacloprido, tebuconazol, triadimefom e triadimenol, garantindo a eficiência da metodologia.

O fungo entomopatogênico Beauveria felina produz as destruxinas, hexadepsipeptídeos cíclicos que apresentam diversas atividades biológicas, como por exemplo: atividade inseticida, fitotóxica, citotóxica contra células tumorais, atividade antiviral contra o vírus da hepatite B, dentre outras. Devido à atividade inseticida das destruxinas, os fungos que as produzem têm grande importância econômica. Assim, o objetivo deste trabalho foi realizar a otimização das condições de crescimento e produção das destruxinas pelo fungo Beauveria felina utilizando diferentes meios de cultivo, a fim de se isolar destruxinas em maiores quantidades para a avaliação de seu potencial tóxico. Foram utilizados métodos de planejamento experimental e análise multivariada para a otimização das condições de crescimento, resultando em três condições ótimas de cultivo para cada um dos meios de cultura MF, PDB e SBD. Em todas as condições ótimas de cultivo foram observadas destruxinas conhecidas, desconhecidas e destruxinas que não foram isoladas anteriormente de Beauveria felina. O cultivo de Beauveria felina no meio PDB foi o que mais apresentou destruxinas desconhecidas (5). Em seguida foi realizada uma otimização das condições de análise cromatográfica de destruxinas por CLAD-DAD-DELE-EM e a validação deste método. Uma das frações obtidas do meio de cultura de Beauveria felina apresentou atividade vermífuga contra Haemonchus contortus. Duas de suas frações purificadas, P1 (destruxina Ed1 - m/z 625 e pseudodestruxina B ou pseudodestruxina C - isobáricas, m/z 669) e P7 (destruxina D ou hidroxihomodestruxina B ou roseotoxina C - isobáricas, m/z 623), foram ativas contra o carrapato bovino Rhipicephalus microplus. / The entomopathogenic fungus Beauveria felina produces destruxins, cyclic hexadepsipeptides which have several biological activities, such as insecticidal, phytotoxic, cytotoxic against tumor cells, antiviral activity against hepatitis B virus, among others. Because of the insecticidal activity of destruxins, fungi that produce them have a considerable economic importance. Therefore the aim of this study was to optimize the conditions for the growth of Beauveria felina and its production of destruxins by using different culture media, in order to isolate larger quantities for the evaluation of their toxic potential. Experimental design and multivariate analysis were used for the optimization of growth conditions, resulting in three conditions for optimum growth for each of the culture media MF, PDB and SBD. Both known and unknown destruxins were observed in each of the optimal conditions, along with destruxins that were not previously isolated from Beauveria felina. The cultivation of Beauveria felina in PDB was the one that showed the largest amount of unknown destruxins (5). We then carried out an optimization of the chromatographic analysis of destruxins by HPLC-PDA-ELSD-MS and the validation of this method. One of the fractions obtained from the culture medium of Beauveria felina showed anthelmintic activity against Haemonchus contortus, while two of its purified fractions, P1 (destruxin Ed1 - m/z 625 and pseudodestruxin B or pseudodestruxin C - isobaric m/z 669) and P7 (destruxin D or hydroxyhomodestruxin B or roseotoxin C - isobaric m/z 623), were active against the cattle tick Rhipicephalus microplus.

This thesis was made possible by the company Verendus system AB, which is the leading developer of dealership management systems in the caravan and camper market. The goal of the thesis was to create a statistics module for a new producer system envisioned by Verendus. The new system is aimed at manufacturers and importers of caravans and campers, with the goal of enabling its users to create more effective production lines. By providing statistical data to mentioned users they will be able to predict market trends and customize their production lines accordingly. Today such software does not exist which leads Verendus to think that its arrival would lead to a success on the market. Before the development and design of the producer system began, a new agile method had to be developed, MAM (Minimum Agile Method). The method has strong influences from already established agile methods such as Scrum, Extreme Programming (XP) and Disciplined Agile Delivery (DAD) and was used during the development of the producer system. The development of the basic statistics module resulted in an administration panel where new users and present users can be added or edited. The administrator can also create new connections and permissions between users and companies, which leads to specific information being displayed for each user. The major part of the development resulted in a page for viewing the statistical data currently present in the database of Verendus. With the provided statistics the user can then make qualified decisions and forecasts that have the potential of directly influencing sales. / Arbetet har ägt rum på företaget Verendus System AB, som är ledande affärssystemsutvecklare inom husvagns- och husbilsmarknaden. Målet med examensarbetet var att skapa en grundmodul för att presentera statistik för husvagns- och husbilstillverkare och importörer. Detta för att de ska kunna effektivisera produktionen och spå trender i ett tidigt stadie. En sådan tjänst existerar inte idag, varpå produkten tros ha stor effekt på marknaden. Innan utveckling- och designarbetet av leverantörsstödet började, skapades en agil arbetsmetod, MAM (Minimum Agile Method). Metoden har starka influenser från redan etablerade agila metoder som Scrum, XP och DAD. MAM applicerades sedan på examensarbetets utvecklingsdel. Leverantörsstödets grundmodul resulterade i en administratörspanel, en inställningssida och en statistiksida. Administratörspanelen används för att kontrollera nya användares behörigheter. Inställningssidan ger användaren möjligheten att ändra sina personuppgifter och statistiksidan är till för att användaren ska kunna generera statistik efter ett antal parametrar, fatta kvalificerade beslut och spå kommande trender.

The aim of this diploma thesis was to determine the effect of the addition of several types of local wines in the production of wine sausages. Selected wine varieties were characterized using basic and modern analytical methods (liquid chromatography with a diode array detector) and the obtained results were compared with the results of sensory analysis.The sensory analysis was divided into two parts, the first evaluated six non-traditional wine varieties used in the production of wine sausages and the second part evaluated samples with the addition of the traditionally used wine variety Veltlínské zelené, but in three different volumes. Using the Statistica program, the data obtained from the sensory analysis were processed and the mutual correlation was evaluated. Using the gas chromatography method with a flame ionization detector the fatty acid content of a standard sample of wine sausage has been determined. Based on all the determined results, the best wine variety for the production of wine sausages was chosen, it was the blue grape variety André.

Content: The restriction of certain dangerous substances according to REACH (Registration, Evaluation, Authorisation of Chemicals as well as the Restricted Substances Lists (RSL) requirements promoted by various renowned brands obliges tanneries to everyday more numerous analysis with undesired conflictual situations on false positive tests results. This situation is worsened by the voluntary reduction of requested detection limits far below the levels recommended by the accepted ISO methods. On this context, ISO 17070:2015 was extended in its current version from the determination of pentachlorophenol to tetrachlorophenol, trichlorophenol, dichlorophenol and monochlorophenol isomers. Some brands also included under this technique the ortho phenyl phenol (OPP) analysis, requesting a quantification limit below 1 mg/kg for all analytes. The present paper proposes a new HPLC-DAD-MS direct method for the verification of chlorophenols positive cases, and its extension to leather chemicals analysis, as well as the discrimination among false positive cases and real positive ones. The paper also illustrates case studies reporting differences in the quantification of the said analytes and the chlorophenol scission in different analytical conditions. Take-Away: False positives verification of the presence of chlorophenols. Quicker method respect to the current ISO 17070:2015 standard. Method reliable on Leather chemicls chlorophenos research. Generation of chlorophenols from certain AOX substances.

This diploma thesis focuses on optimization and validation of the method for a mixture of tetracycline antibiotics (oxytetracycline, tetracycline) determination using high performance liquid chromatography with a diode array detector (HPLC–DAD). The aim of this thesis was to determine suitability of the system for the determination of these antibiotics in milk. The theoretical part is focused on individual groups of antibiotics and their mechanism of action, resistances and possibilities of use. Next point of this thesis is legislation, which describes permitted amounts of antibiotics in milk. In this study, the HPLC method and validation parameter is described in detail. The experimental part deals with creation and optimization of the method and its validation. Following validation parameters were verified: system suitability, linearity, method accuracy, limit of detection and quantification, robustness. The results obtained indicate mostly suitable ratio of mobile phase of 0,01 M trifluoracetic acid:methanol:acetonitrile = 80:10:10. The wavelength 355 nm was found as the most feasible for this method after evaluating the result. Determination of robustness showed that the method is robust, and the only significant effect on the robustness was observed in column changes.

In der vorliegenden Arbeit wurde die Leistungsfähigkeit der Hochleistungsflüssigchromatographie mit Photodiodenarraydetektion (HPLC-DAD) im Rahmen der Systematischen Toxikologischen Analyse (STA) umfassend untersucht, ein besonderer Schwerpunkt lag dabei auf der Aussagesicherheit bei der Substanzidentifizierung. Die Grundlage für diese Untersuchungen bildete die Messung der UV-Spektren und relativen Retentionszeiten (RRT) von 2.682 toxikologisch relevanten Substanzen mit Hilfe moderner Diodenarraydetektoren und unter einheitlichen experimentellen Bedingungen (Eluent: Acetonitril/ Phosphatpuffer pH 2,3; Trennsäule: Lichrospher RP8ec). Die erhaltenen Daten wurden zusammen mit weiteren Substanzinformationen in einer Spektrenbibliothek zusammengefasst, die als Buch und CD-ROM auch der Öffentlichkeit zugänglich gemacht wurde. Bei der Entwicklung eines Konzeptes zur Methodenvalidierung für die STA erwiesen sich insbesondere die Kenngrößen Selektivität und Spezifität als von zentraler Bedeutung. Unter Verwendung eines Fensteransatzes für die RRT und des Similarity Index (SI) für den Spektrenvergleich wurden sämtliche möglichen Substanzpaare aus der Spektrenbibliothek miteinander auf ihre Unterscheidbarkeit hin verglichen, wobei verschiedene RRT-Fensterbreiten und SI-Schwellenwerte zugrundegelegt wurden. Aus den Ergebnissen wurden Selektivitätsparameter wie die Relative Identification Power (RIP), die Discriminating Power (DP) und die Mean List Length (MLL) berechnet. Mit einer RIP von 84,2 %, einer MLL von 1,25 und einer DP von 0,9999 wurden für die Substanzidentifikation anhand von RRT + UV-Spektrum im Literaturvergleich sehr gute Ergebnisse erhalten. Die hohe Grad an Spezifität der UV-Spektren für den Einsatz in der STA wurde einerseits durch Auszählung sämtlicher in der Spektrenbibliothek gefundener Chromophore/Absorptionssysteme belegt: 1.650 verschiedene Absorptionssysteme wurden gefunden, über 60 % der Verbindungen trugen ein in diesem Substanzbestand einzigartiges Absorptionssystem. Andererseits wurde die computergestützte Unterscheidbarkeit zwischen verschiedenen Gruppen jeweils strukturell ähnlicher Wirkstoffe anhand des UV-Spektrums untersucht. Hierfür wurden 487 Substanzen in ein hierarchisches System von Haupt- und Untergruppen eingeteilt, zwischen denen anschließend mit Hilfe der Schrittweisen Diskriminanzanalyse (SDA) unterschieden werden sollte. Die sehr guten Klassifizierungsergebnisse und die Auflösung auch feiner struktureller Unterschiede, oftmals abseits des eigentlichen Absorptionssystems, lassen es grundsätzlich möglich erscheinen, auch Substanzen für die STA zu erschließen, die nicht in der Spektrenbibliothek enthalten sind. Insgesamt erwies sich die HPLC-DAD als hochselektives und -spezifisches Messverfahren, das zu Recht eine der wichtigsten STA-Methoden darstellt. / In this thesis the capability of high-performance liquid chromatography with photodiode array detection in the context of Systematical Toxicological Analysis (STA) was extensively investigated with a special focus on the certainty of substance identification. As the foundation for these investigations, UV spectra and relative retention times (RRT) of 2,682 toxicologically relevant compounds were recorded under uniform experimental conditions (mobile phase: acetonitrile/ phosphate buffer pH 2.3; column: Lichrospher RP8ec) using up-to-date photodiode array detectors. Together with additional substance information these data were built into a spectra library also made available to the public in the form of a book and a CD-ROM. During the development of a concept for the validation of a STA method, selectivity and specificity were found to be of central importance. Using a window approach for the RRT and the Similarity Index (SI) for spectral comparison, all possible substance pairs within the spectra library were examined in terms of their respective distinguishability, based on different RRT windows and SI threshold values. From the results selectivity parameters such as the Relative Identification Power (RIP), the Discriminating Power (DP), and the Mean List Length (MLL) were calculated. As compared to the literature, with a RIP of 84.2 %, a MLL of 1.25, and a DP of 0,9999 excellent results were obtained for the substance identification based on the combination of RRT and UV spectrum. The high degree of specificty of UV spectra for use in the STA was demonstrated on the one hand by counting all different chromophores/ absorption systems found in the spectra library: 1,650 systems were found, more than 60 % of the substances bore absorption systems unique in the dataset. On the other hand the computer-based distinction between different groups of structurally similar compounds on the basis of the UV spectrum was investigated by dividing 487 substances into a hierarchical system of main and subgroups and trying to distinguish between these groups using stepwise discriminant analysis (SDA). The highly satisfying classification results together with the resolution of fine structural differences, often remote from the actual absorption system, make it seem possible to also render substances not included in the spectra library accessible for the STA. In conclusion HPLC-DAD proved to be highly selective and specific, rightfully regarded as one of the most important STA methods.